EMBRYOPROTECTIVE ROLE OF ENDOGENOUS CATALASE By

Oxidative stress and reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), which is detoxified by catalase, are implicated in fetal death and birth defects, but embryonic levels of catalase are only about 5% of adult activity, and its protective role is unknown. Our approach involved the use of mice genetically modified to either: (1) express low levels of endogenous catalase (acatalasemic, aCat); or, (2) express human catalase resulting in elevated levels of embryonic catalase activity (hCat). Using these mouse models we investigated the protective importance of constitutive embryonic catalase against endogenous ROS and the ROS-initiating teratogen phenytoin in embryo culture and in vivo. We hypothesized that aCat mice would be more sensitive to endogenous embryonic and phenytoin-enhanced ROS production, while hCat embryos would be less sensitive. aCat and hCat embryos respectively exhibited reduced and enhanced catalase activity compared to wild-type (WT) controls, with conversely enhanced and reduced spontaneous and phenytoin-enhanced embryopathies and DNA oxidation. Among aCat embryos exposed to phenytoin, embryopathies increased with decreasing catalase activity, and were completely blocked by addition of exogenous catalase. The alterations in phenytoin embryopathies were not due to pharmacokinetic differences, as drug concentrations in maternal and fetal tissues were similar among all strains. However, phenytoin concentrations in fetal brain exceeded those in fetal liver or maternal tissues, which may explain the predominance of cognitive deficits over structural birth defects in children exposed in utero to phenytoin. Similarly in untreated aged mice (about 18 months), female aCat mice showed a substantial loss in motor coordination compared to WT controls in the rotarod test. Following in utero exposure to phenytoin, the effect of altered embryonic catalase activity on postnatal neurodevelopment was assessed by several preand post-weaning tests. Catalase deficiency (aCat), independent of drug treatment, reduced performance in surface righting, negative geotaxis tests and rotarod tests. Conversely, high catalase expression (hCat) enhanced performance in the surface righting, negative geotaxis, air righting and rotarod tests. Our results provide the first evidence that the